The study of circulating tumor cells can yield important biologic and clinical information. Yet, it has been limited by the paucity of specific and sensitive markers. Pre-genomic approaches rely on known tumor-associated phenotypes or genetic aberrations. For example, cytokeratins were widely used as markers of circulating breast cancer cells (Pantel, K., et al. 2004 Nat Rev Cancer 4:448-56).
Molecular-based methods for the detection of cancer have greatly energized the cancer community because of their promise of exquisite specificity and sensitivity (Sidransky, D. 1997 Science 278:1054-9). Nevertheless, measuring DNA from tumor cells by PCR has largely been restricted to known tissue-specific antigens and tumor-associated mutations in oncogenes or in tumor suppressors. RT-PCR has also been used to detect tumor-specific transcripts, but again mRNA sequences were primarily based on known aberrations. Most importantly, it is not uncommon for these so called “tumor-specific” transcripts to be found in normal hematopoietic cells.
Sophisticated use of chemotherapy, surgery, and/or radiation therapy can reduce cancers to near complete remission. However, cancer cure remains elusive, the major hurdle being minimal residual disease (MRD) below the detection limit of conventional radiographic or histopathological tools. Since the current eligibility criteria of most clinical trials require evidence of gross disease, a tumor will not be treated until it is measurable and symptomatic. This “killing paradigm” maybe undesirable for several reasons (Schipper, H., et al. 1995 J Clin Oncol 13(4):801-7). First, the Goldie-Coldman hypothesis predicts that bigger tumors have higher likelihoods of mutations and resistance (Goldie, J. H., et al. 1989 Devita H R, ed. Cancer. Principles and Practice of Oncology. 3rd ed. Philadelphia, Pa.: JP Lippincott. 1-12). Second, visible tumors acquire additional barriers to drug delivery (e.g. suboptimal tumor pressure, vasculature and oxygenation) (Semanza, G. L. 2003 Nat Rev Cancer 3(10):721-32; Jain, R. K. 2005 Science 307(5706):58-62). Third, a patient with measurable tumors is physically and/or mentally compromised, and is less likely to tolerate treatment side effects. An alternative strategy of “regulatory control” has been proposed (Schipper, H., et al. 1995 J Clin Oncol 13(4):801-7). This treatment paradigm is particularly relevant to today's cancer therapeutics. While novel agents such as angiogenesis inhibitors, growth modulators or vaccines may not achieve rapid tumor shrinkage, they may nevertheless be effective in controlling MRD, such that patients can “live with cancer”.
Targeting subclinical disease is particularly pertinent to high risk neuroblastoma (NB), a pediatric cancer which poses enormous clinical challenges because of its tumor bulk, the extent of metastatic spread, and its orphan disease status. Although most patients achieve near complete remission, they typically relapse because of refractory MRD. Adjuvant therapies such as stem cell transplantation and immunotherapy are employed, but the ability to measure MRD accurately is crucial to determine their anti-tumor effect, to identify the optimal timing for stem cell collection, and to provide early indications of treatment failure.
The detection of MRD using molecular-based methods shows great promise because of its exquisite specificity and sensitivity (Sidransky, D. 1997 Science 278(5340): 1054-9). One such technique is quantitative reverse transcription-polymerase chain reaction (qRT-PCR) to measure tumor transcripts. The clinical utility of tyrosine hydroxylase (TH), an established NB marker, has been extensively described (Naito, H., et al. 1991 Eur J Cancer 27:762-765; Miyajima, Y., et al. 1995 Cancer 75:2757-2761). GD2 synthase (β1,4-N-acetylgalactosaminyltransferase, GD2/GM2 synthase, GalNacT) is also a useful MRD marker in the bone marrow (BM) and peripheral blood (PB) among high risk NB patients (Cheung, I. Y., et al. 2001 Clin Cancer Res 7(6):1698-705; Cheung, I. Y., et al. 2003 J Clin Oncol 21(6):1087-93).
In order to prevent relapse, adjuvant therapy such as autologous marrow or peripheral blood stem cell transplantation and immunotherapy are often employed. It stands to reason that accurate quantitation of MRD can identify the optimal timing for stem cell collection and evaluate the efficacy of adjuvant therapies, permitting the monitoring of tumor activity previously undetectable by standard histologic and radiographic techniques. In children, where late effects of prolonged treatment are of grave concern, specific and sensitive markers of MRD may provide objective endpoints for terminating cytotoxic therapy.
Thus, improved methods for the identification of patients having MRD are desired for the diagnosis and treatment of cancer patients.